DNA and RNA encoding proteins useful in the regulation of KB-containing genes, and cells containing same

ABSTRACT

Proteins, and corresponding DNA and RNA sequences, useful for the regulation of expression of κB-containing genes are disclosed. These proteins are useful to either stimulate or inhibit the expression of κB-containing genes. Proteins stimulating the expression of κB-containing genes have an amino acid sequence at least 80% identical to the amino acid sequence of from position 1 to position 374 of p100 [SEQ ID NO: 2]. Proteins having an inhibitory effect on the expression of κB-containing genes have sequences either at least 80% identical to the amino acid sequence of from position 407 to the carboxyl end of p100 [SEQ ID NO: 2] or having an amino acid sequence at least 80% identical to the amino sequence of either from position 1 to 100 sor from position 101 to position 374 of p100 [SEQ ID NO: 2].

This is a division, of application Ser. No. 07/747,781, filed on Aug.20, 1991, now U.S. Pat. No. 5,324,818.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to genetic sequences and proteins usefulin the regulation of genes, particularly in the regulation ofκB-containing genes.

2. Discussion of the Background

A twice-repeated 11-base-pair sequence, termed κB, which is known toexist in association with the enhancer of the immunoglobulin κ lightchain (Sen et al, Cell (1986) 46:705, Sen et al, Cell (1986) 47:729) ispresent in the control elements of numerous genes, including both viraland cellular genes. NF-κB, a regulatory element originally described asa transcription factor that recognizes the κB element of theimmunoglobulin (Ig) κ light chain enhancer and which is known to beassociated with κ light chain expression, has been found in a variety ofviral enhancers, including SV40, CMV, HIV-2 and SIV. κB-related siteshave been found in association with several cellular genes, includingclass I and II MHC genes, IL-2, IL-2 receptor (IL-2Rα), serum amyloid A,β-2 microglobulin, β-interferon, tumor necrosis factor, and T-cellreceptor genes.

A κB-like site contributes functionally to IL-2-dependent geneexpression and may also regulate IL-2Rα transcription. κB is also foundamong the regulatory elements upstream of the HIV enhancer. This sitewas initially identified as a positive regulatory element recognized byDNA binding proteins found to be present in activated, but not resting,T cells. Mutation of the κB sites abolished inducibility of the HIVenhancer in T leukemia cells. A link between the κB site and the tat-Igene was also suggested by the synergistic response of the HIV enhancerto tat-I and NF-κB-mediated stimulation.

Another κB-like site was found upstream of the class I MHC gene.Although this site competes for binding to NF-κB, a DNA binding proteindistinct from NF-κB has been identified in MEL and HeLa cells. Baldwinet al, Proc. Nat. Acad. Sci. (USA) (1988) 85:723. This protein, termedH2TF1, was detected in nuclear extracts containing no detectable NF-κBbinding activity and had a different apparent molecular weight asmeasured by UV-cross-linking analysis. Another protein, designated KBF1,also recognized this site and may be related in part to NF-κB. Israel etal, Proc. Nat. Acad. Sci. (USA) (1987) 84:2653, Yanno et al, EMBO J(1987) 6:3317.

Although these proteins recognize κB-like sites, their relationship tothe IL-2R κB binding protein(s) and NF-κB is unknown. Similarly, theHIVen86 protein is likely to be distinct from NF-κB since it has ahigher apparent molecular weight. Franza et al, Nature (1987) 330:391.Its relationship to the IL-2R κB binding protein, which has beendesignated RκB (NF-rκB), and its role in mediating transactivationdependent on these κB-like sites is also not yet understood. Althoughthe evidence is indirect, it appears that HIVen86 also differs from RκB,which has a higher apparent molecular weight of 100 kD.

While extensive sequence similarity to κB is found among enhancersassociated with several genes (Lung et al, Nature (1988) 333:776), asecond characteristic shared by these sites is their ability to respondto transactivation by the tax₁ gene of HTLV-1. At the same time, severallines of evidence suggest differences among κB binding proteins. Forexample, nuclear extracts from MEL cells display an H2TF1 bindingactivity in the absence of detectable NF-κB binding. Competition studiesshowed that both the IL-2RκB and κB sites compete less efficiently thanH2TF1 for binding to the site, consistent with previous studiessuggesting that a distinct binding protein binds to the H2TF1 site.

Analysis of binding proteins by UV-cross-linking and SDS-page has alsorevealed multiple radiolabeled complexes, further implicating multipleproteins in κB binding. By UV-cross-linking, several specific complexeshave been detected, including proteins of molecular weights of ca. 160,ca. 90, ca. 75, and ca. 50 kD. There is no evidence for differentialregulation of these proteins by different NF-κB stimulants since PMA,TNF-α, and IL-1 induce the same set of complexes. The complex of ca. 50kD is consistent with previous report of NF-κB and/or KBF1. Although aκB binding protein of 86 kD HIVen86A, has also been identified bytwo-dimensional gel electrophoresis, it is unclear whether the ca. 90 kDprotein represents this protein. Both HIVen86 and RκB show no increasein binding following cellular activation. The 160 kD complex has notbeen previously described.

In summary, at least seven κB binding proteins have been defined to dateby either mobility shift analysis, UV-cross-linking, or proteinpurification. The molecular weight of these proteins range in size from48 to greater than 300 kD. The various κB binding proteins that havebeen reported are indicated in Table 1 below, together with theirrelative specificities for the canonical κB, the class I MHC, and theIL-2RκB sites.

                  TABLE 1                                                         ______________________________________                                        Summary of κB Binding Proteins                                                                     Protein  CDNA                                      Name       Specificity     (kD)     (kb)                                      ______________________________________                                        NF-κB/KBFI                                                                         κB = MHC = IL-2RκB                                                                50       4                                         H2TF-1     MHC > κB > IL-2RκB                                                                110      --                                        EBP-L      MHC = κB  60       --                                        HIVen86    KB = IL-2RκB                                                                            86       --                                        MBP-1      MHC > κB > IL-2RκB                                                                ˜300                                                                             9.5                                       RκB (IL-2RκB)                                                                IL-2RκB > κB > MHC                                                                95       5.5                                       ______________________________________                                    

Of these, the cDNAs encoding three of these proteins have been isolated.These cDNAs differ from one another by primary sequence and react withmRNAs of distinct size by northern blot. Taken together, these dataindicate that multiple proteins could bind to a set of κB-related sites.Thus, there is a family of κB binding proteins, which, unlike some DNAbinding proteins (e.g., c-jun), do not appear to be members of a relatedmultigene family.

The κB sequence is known to exist in association with the regulatoryelements of various viral and cellular genes. In light of the obviousinterest in regulating gene expression there is accordingly a need for afactor useful in the regulation of κB-containing genes.

SUMMARY OF THE INVENTION

Accordingly, it is an object of this invention to provide novel factors,and materials and methods useful for obtaining and using such factors,useful in the regulation of κB-containing genes.

The above objects, of the invention, and other objects which will becomeapparent from the description of the invention given hereinbelow, havebeen surprisingly found to be satisfied by proteins having amino acidsequences related to p49 [SEQ ID NO: 1] and p100 [SEQ ID NO: 2]. Moreparticularly, the inventors have discovered that proteins having aminoacid sequences at least 80%, preferably at least 90% and more preferablyat least 95%, identical to either (i) the sequence of from amino acidposition about 1 to amino acid position about 374 and up to the completesequence of p49 [SEQ ID NO: 1] or (ii) the sequence of from amino acidposition about 1 to amino acid position about 374 and up to amino acidposition about 500 of p100 [SEQ ID NO: 2] are useful to stimulate theexpression of κB-containing genes. The inventors have also discoveredrelated proteins useful in inhibiting the expression of κB-containinggenes. These proteins may be characterized by amino acid sequences atleast 80%, preferably at least 90% and more preferably at least 95%,identical to the sequence of from amino acid position about 407 of p100[SEQ ID NO: 2]. The proteins exhibiting the inhibitory effect may alsobe characterized by amino acid sequences at least 80%, preferably atleast 90% and more preferably at least 95%, identical to the sequence offrom either (i) amino acid positions about 1 to about 190 or (ii)positions about 191 to about 374 of p100 [SEQ ID NO: 2] and up to aboutcomplete p100 [SEQ ID NO: 2].

BRIEF DESCRIPTION OF THE FIGURES

The figures set forth the deduced amino acid sequence of the p49 [SEQ IDNO: 1] and p100 [SEQ ID NO: 2] NF-κB proteins--two proteins provided bythe invention--and their similarity to other NF-κB/rel/dorsal genes.More particularly, FIG. 1 provides a schematic comparison of the majorstructural features of p49 [SEQ ID NO: 1] and p100 [SEQ ID NO: 2]. FIG.2 provides the deduced amino acid sequence of p49 [SEQ ID NO: 1] andp100 [SEQ ID NO: 2] and comparison to p105 NF-κB [SEQ ID NO: 3].

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The transcription factor NF-κB is a protein complex which comprises aDNA-binding subunit and an associated transactivation protein (ofrelative molecular masses 50,000 (50K) and 65K, respectively). Both the50K and 65K subunits have similarity with the rel oncogene and theDrosophila maternal effect gene dorsal. The 50K DNA-binding subunit waspreviously thought to be a unique protein, derived from the 105K geneproduct (p105). The present invention is based on the inventor'sdiscovery of a complementary DNA (cDNA) that encodes an alternativeDNA-binding subunit of NF-κB. This cDNA is more similar to p105 NF-κBthan other family members and defines a new subset of rel-related genes.

It is synthesized as a ca.100K protein (p100) that is expressed indifferent cell types, contains cell cycle motifs and, like p105, must beprocessed to generate a 50K form. A 49K product (p49) can be generatedindependently from an alternatively spliced transcript; it has specificκB DNA-binding activity and can form heterodimers with other relproteins. In contrast to the ca.50K protein derived from p105, p49 actsin synergy with p65 to stimulate the human immunodeficiency virus (HIV)enhancer in transiently transfected Jurkat cells. The p49/p100 NF-κB istherefore important in the regulation of κB-containing genes, includingHIV.

The present invention accordingly provides proteins useful in theregulation of κB-containing genes. These proteins may be used to eitherstimulate or inhibit the expression of κB-containing genes. Theseproteins are based on the inventor's discovery that the amino acidsequence located at positions about 1 to about 374 of p49 [SEQ ID NO: 1]and up to the complete sequence of p49 [SEQ ID NO: 1], simulates theexpression of κB-containing genes. (Note that p49 [SEQ ID NO: 1] andp100 [SEQ ID NO: 2] are identical from positions 1 to 374.)

The inventor has further discovered that the amino acid sequence foundat about positions about 1 to about 190 of p100 [SEQ ID NO: 2] is a DNAbinding domain and that the amino acid sequence found at positions about191 to about 374 of p100 [SEQ ID NO: 2] is a dimerization interface. Andthe inventor has still further discovered that the amino acid sequencefound at position about 407 of p100 [SEQ ID NO: 2] going all the way toabout the carboxyl terminal of the protein is also capable of inhibitingthe expression of κB-containing genes.

The proteins provided by the present invention which accordingly act tostimulate κB-containing genes have an amino acid sequence at least 80%identical (i.e. up to 20% of the amino acids have been changed, deleted,or both), preferably 90% identical, and most preferably 95% identical,to the amino acid sequence of amino acid positions about 1 to about 374of either p49 [SEQ ID NO: 1] or p100 [SEQ ID NO: 2] and optionally up toabout the complete sequence of p49 [SEQ ID NO: 1] or up to amino acidposition about 500 of p100 [SEQ ID NO: 2].

Proteins useful in inhibiting κB-containing genes provided by theinvention are of two related types. The first is a transdominant mutantof p49 [SEQ ID NO: 1] in which either (i) the dimerization interfaceregion of p49 [SEQ ID NO: 1] has been retained (located at positionsabout 191 to about 374 of p100 [SEQ ID NO: 2]) or (ii) the DNA bindingregion (located in positions about 1 to about 190 of p100 [SEQ ID NO:2]) has been retained. In either of these two embodiments the other(i.e. unretained) region has been either removed or altered to causeloss of DNA binding or of dimerization activity, e.g., by removing orreplacing at least 10% of its amino acids, preferably at least 25% ofits amino acids, to achieve removal of DNA binding or dimerizationactivity.

This transdominant mutant of p49 [SEQ ID NO: 2] may be a protein havingan amino acid sequence at least 80% identical, preferably 90% identical,and most preferably 95% identical, to either (i) the amino acid sequenceof amino acid positions about 1 to about 190 or (ii) the amino acidsequence of amino acid positions about 191 to about 374 of either p49 orp100 [SEQ ID NO: 1 or 2] and optionally up to the complete p49 [SEQ IDNO: 1] or complete p100 [SEQ ID NO: 2]. That is, if the protein is onein which the DNA binding region has been altered or removed, the proteinmay further comprise, attached to its carboxyl end, an amino acidsequence at least 80% identical, preferably 90% identical, and morepreferably 95% identical to up to 100%, the amino acid sequence of frompositions 374 to the carboxyl end of p100 [SEQ ID NO: 2].

With the above transdominant mutant of p49 [SEQ ID NO: 1] one useseither the protein segment comprising the DNA binding region or theprotein segment comprising the dimerization interface region of p49 [SEQID NO: 1]. The DNA binding region may be used by itself or it may beused in association with an inactivated dimerization interface, wherethe inactivation results from removal of at least 10%, or preferably 25%amino acids from the dimerization interface region. If a protein segmentcorresponding to the dimerization interface region is used, it may beused by itself or in association with an inactivated DNA binding region,where inactivation is caused by the removal of at least 10%, preferablyat least 25% of the amino acids from the DNA binding region.

Another group of κB inhibiting proteins provided by the presentinvention have an amino acid sequence at least 80% identical, preferably90% identical, and most preferably 95% identical to the sequence ofamino acid at positions about 407 of p100 [SEQ ID NO: 2] to about thecarboxy end of p100 [SEQ ID NO: 2].

The present invention also provides DNA and RNA sequences encoding tothe above proteins, corresponding antisense RNA sequences, vectorsuseful to express these proteins, and both eukaryotic and prokaryoticcells containing these DNA and/or RNA sequences, further optionallycontaining a κB-containing gene.

The antisense RNA sequences provided by the present invention may be oneof five different types. The first is an antisense RNA sequence which isat least 20 nucleotides-long and corresponds to an amino acid sequencefalling within positions 1 to 374 of p49 [SEQ ID NO: 1], inclusive. Thesecond is an antisense RNA sequence which is at least 20nucleotides-long and corresponds to an amino acid sequence fallingwithin the sequence of p49 [SEQ ID NO: 1]. The third is an antisense RNAsequence which is at least 20 nucleotides-long and corresponds to anamino acid sequence falling within the sequence of p100 [SEQ ID NO: 2].The fourth is an antisense RNA sequence which is at least 20nucleotides-long which corresponds to an amino acid sequence fallingwithin the sequence of from position 375 to the carboxyl end of p49 [SEQID NO: 1], inclusive. The fifth is an antisense RNA sequence which is atleast 20 nucleotides-long and corresponds to an amino acid sequencefalling within positions 375 to the carboxyl end of p100 [SEQ ID NO: 2],inclusive.

These antisense RNA sequences may be used in accordance with knowntechniques (see, e.g., Zamecnik et al., Proc. Nat. Acad. Sci. (USA),(1986) 83:4143-4146) to inhibit or block cellular expression of thegenes encoding either p49 [SEQ ID NO: 1] or p100 [SEQ ID NO: 2]. Moreparticularly the above first, second and third antisense RNA sequencesmay be used to inhibit or block expression of either p49 [SEQ ID NO: 1]or p100 [SEQ ID NO: 2]. The fourth antisense RNA sequence may be used toinhibit or block expression of p49 [SEQ ID NO: 1] because it is drawn toa sequence unique to p49 [SEQ ID NO: 1]. The above fifth antisense RNAsequence may be used to inhibit or block expression of p100 [SEQ ID NO:2] because it is drawn to a sequence unique to p100 [SEQ ID NO: 2].

The eukaryotic and prokaryotic cells provided by the present inventioncontain the DNA and/or RNA sequences encoding a present protein,optionally together with a κB-containing gene. These latter sequences(i.e., the sequences containing the κB sites) are present in these cellsin a geometry permitting regulation of the κB-containing gene by theproteins of the present invention.

The p49/p100 subunit of NF-κB provided by the invention can be used toregulate the expression of recombinant genes in eukaryotic orprokaryotic cells. In one form of this method, the p49/p100 gene can beoverexpressed in a host cell which contains a separate κB-dependentenhancer to express a given recombinant gene. When the p49 gene isoverexpressed, the κB-regulated enhancer will stimulate expression ofthe recombinant gene of interest.

In another method, it is possible to use the κB site to blocktranscriptional activation. By placing the κB binding sites within apromoter near the transcriptional initiation site, it would interferewith transcriptional initiation or elongation when the recombinantp49/p100 proteins could bind to these sites. It is therefore possible toinhibit expression of a specific recombinant gene when this molecule isoverexpressed.

Another way to regulate expression of recombinant genes within cells isto make a fusion protein between p49 (or p100) gene products, forexample, the DNA binding and dimerization domains and fuse them to anacidic transactivation domain, such as that of the herpesvirustransactivator, VP16. Such chimeric constructs would provide a highlevel of constitutive transactivation to κB-dependent plasmids.

Several such plasmids can be synthesized whereby fusion proteins havebeen linked to the 3'-end of either p49 or an equivalent of p100,creating potent transactivators. Such chimeric transactivators can alsobe used to overexpress recombinant proteins with cells.

The NF-κB-related DNA and RNA or protein sequences provided by theinventors can be utilized for a variety of different functions. First,the recombinant protein can be used to generate antibodies against thisgene product to detect this protein within cells or tissues. Since theseproteins are involved in the regulation of HIV and may also be relatedto cell division and malignancy, this would provide a useful assay todetermine the degree of progression of HIV or the degree of activationof malignant cells.

In addition, the DNA or RNA sequences can be measured directly followingHIV infection as an indicator of the activity of virus within cells. Inrecent studies, the inventors have noted a decrease in p100 expressionfollowing HIV infection in cell culture. The recombinant proteins canalso be used to generate transdominant mutants which can be used toinhibit activation of HIV, normal cellular genes, or cell replication.The purified recombinant proteins may also be used to define mechanismsof transcriptional activation in the laboratory to determine thespecificity of DNA binding, and to determine the 3-dimensional structureof these proteins for further attempts at molecular modeling andrational drug design.

Cultures of E. coli XL-blue cells transformed with bluescript plasmidscontaining p49 [SEQ ID NO: 1] or p100 [SEQ ID NO: 2] encoding insertshave been deposited on Aug. 20, 1991 in the permanent culture collectionof the American Type Culture Collection (ATCC), located at 12301Parklawn Drive, Rockville, Md. 20852 USA. These cultures have beenaccorded the following accession numbers: ATCC68673 and ATCC68672.

Having generally described this invention, a further understanding canbe obtained by reference to certain specific examples which are providedherein for purposes of illustration only and are not intended to belimiting unless otherwise specified.

EXPERIMENTAL

To characterize the molecular structure of NF-κB and its mechanism ofactivation, the inventor attempted to identify cDNAs that encodeproteins within this complex. Trypsin digested κB binding proteinspurified from bovine spleen were sequenced. Three peptides so obtainedwere identical to a DNA binding subunit of human NF-κB/KBF1, whileanother was 75% identical to a homologous peptide of p105. This peptidesequence suggested the existence of an alternative NF-κB protein. A PCRprobe amplified from degenerate primers was found to be identical to a700 bp fragment of human c-rel and was used for low stringencyhybridization.

From 6×10⁵ recombinants, two clones were identified which hybridized atlow stringency. By Southern blot analysis at high stringency, this genewas present in single copy, and Northern blot analysis revealed at leasttwo hybridizable RNA species of ˜1.9 kb and ˜3.5 kb, demonstrating thatmultiple species of p49 existed, and their relative abundance variedamong different cell types. For example, the larger 3.5 kb species,found in all cells examined, was the predominant form in YT T leukemiacells; but both transcripts were present equally in the JY B cell line.

To characterize the larger mRNA species, additional cDNA clones wereisolated and sequenced. The clone for the shorter transcript contained a1341 bp open reading frame, predicting a protein of MW ˜49,100. Thelonger clone encoded a protein of predicted MW, ˜100,634 kd. Thesesequences were identical through amino acid 374, into the glycine-richputative hinge region, after which they diverged. The amino acidsequence in the common N-terminal region was similar (26% identity) toNF-κB, dorsal and rel, with greatest similarity between p49/p100 andp105 (60% identity). One subregion contained conserved cysteine andhistidine residues that do not form a classic zinc finger structure.Interestingly, κB-binding activity is dependent upon zinc, and,analogous to the tat-I gene of HIV, may form an alternative structurewhich participates in dimerization or nucleic acid binding. The longertranscript contained repeated sequences in the C-terminal region withhomology to motifs in p105 and cell cycle genes. Among these familymembers, p100 NF-κB is most closely related to p105 NF-κB (41%identity), while p65 is most similar to c-rel (50% identity).

To characterize the DNA binding activity of the p49 cDNA, theelectrophoretic mobility shift assay (EMSA) was performed using aprokaryotic expression system. The predominant product of ˜49 kddisplayed specific κB binding activity which was competed by the H-2 andHIV κB sites, but not by a single base pair mutant of HIV or anunrelated IL-2 octamer site. Interaction of p49 with other rel familymembers was examined by immunoprecipitation in a wheat germco-translation system similar to p105(Xba I), p49 associated with c-rel.

To determine whether p49 interacts with other NF-κB/rel proteins tostimulate transcription, eukaryotic expression vectors were transfectedinto Jurkat cells. Transfection of p49 alone stimulated κB enhanceractivity slightly, while p65 at higher concentrations significantlyincreased κB-dependent transcription. Transfection of low amounts (≦1μg) of either p49 or p65 caused minimal stimulation, but whenco-transfected together, they acted in synergy to stimulate a κBreporter plasmid. This stimulation was more effective than thecombination of "p50" [p105(Rsa I)] and p65, suggesting that p49 was moreeffective in cooperating with p65. When analyzed with the HIV-CATreporter, the p49/p65 combination, but not p50 [p105(Rsa I)]/p65,stimulated HIV-CAT activity, and the effect required an intact κBregulatory element. A truncated p100 (48.5 kd form) showed similarstimulation, in contrast to full-length p100 which was inactive,suggesting that differences in the rel-conserved domain mediate thiseffect.

These findings demonstrate that κB-dependent transcription is regulatedby the p49/100 gene products. p65 and rel are putative transcriptionalactivation subunits with intrinsic DNA binding activity which associatewith another DNA binding subunit, previously thought to be a single geneproduct derived from p105. These findings suggest that p49/100represents an alternative DNA binding subunit of NF-κB which synergizeseffectively with p65 to activate κB-dependent transcription. Manyproteins can bind to κB-related sites, some of which are notNF-κB/rel-related. Although a variety of κB-binding proteins have beendefined biochemically, their identification has remained equivocal sincerelated antigenic epitopes are contained in their amino terminal region.As with p49/100, the identification of these cDNAs allows moredefinitive analysis of their expression and function. The interaction ofp49/100 with other proteins and its potential alternative modes ofregulation may provide additional mechanisms to regulate thetranscription of HIV and different κB-containing genes.

Obviously, numerous modifications and variations of the presentinvention are possible in light of the above teachings. It is thereforeto be understood that within the scope of the appended claims, theinvention may be practiced otherwise than as specifically describedherein.

    __________________________________________________________________________    SEQUENCE LISTING                                                              (1) GENERAL INFORMATION:                                                      (iii) NUMBER OF SEQUENCES: 3                                                  (2) INFORMATION FOR SEQ ID NO:1:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 447 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: unknown                                                         (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:                                       MetGluSe rCysTyrAsnProGlyLeuAspGlyIleIleGluTyrAsp                             151015                                                                        AspPheLysLeuAsnSerSerIleValGluProLysGluProAlaPro                               202530                                                                       GluThrAlaAspGlyProTyrLeuValIleValGluGlnProLysGln                              354045                                                                        ArgGly PheArgPheArgTyrGlyCysGluGlyProSerHisGlyGly                             505560                                                                        LeuProGlyAlaSerSerGluLysGlyArgLysThrTyrProThrVal                              65 707580                                                                     LysIleCysAsnTyrGluGlyProAlaLysIleGluValAspLeuVal                              859095                                                                        Th rHisSerAspProProArgAlaHisAlaHisSerLeuValGlyLys                             100105110                                                                     GlnCysSerGluLeuGlyIleCysAlaValSerValGlyProLysAsp                               115120125                                                                    MetThrAlaGlnPheAsnAsnLeuGlyValLeuHisValThrLysLys                              130135140                                                                     AsnMetM etGlyThrMetIleGlnLysLeuGlnArgGlnArgLeuArg                             145150155160                                                                  SerArgProGlnGlyLeuThrGluAlaGluGlnArgGluLeuGluGln                               165170175                                                                    GluAlaLysGluLeuLysLysValMetAspLeuSerIleValArgLeu                              180185190                                                                     ArgPheSerAlaPheLeuArgAlaSerAspGlySerPheSerLeuPro                              195200205                                                                     LeuLysProValThrSerGlnProIleHisAspSerLysSerProG ly                             210215220                                                                     AlaSerAsnLeuLysIleSerArgMetAspLysThrAlaGlySerVal                              225230235240                                                                  ArgGlyGlyAspGluValTyrLeuLeuCysAspLysValGlnLysAsp                              245250255                                                                     AspIleGluValArgPheTyrGluAspAspGluAsnGly TrpGlnAla                             260265270                                                                     PheGlyAspPheSerProThrAspValHisLysGlnTyrAlaIleVal                              275280 285                                                                    PheArgThrProProTyrHisLysMetLysIleGluArgProValThr                              290295300                                                                     ValPheLeuGlnLeuLysArgLysArgGlyGlyAspValSerAsp Ser                             305310315320                                                                  LysGlnPheThrTyrTyrProLeuValGluAspLysGluGluValGln                              325330 335                                                                    ArgLysArgArgLysAlaLeuProThrPheSerGlnProPheGlyGly                              340345350                                                                     GlySerHisMetGlyGlyGlySerGlyGlyAl aAlaGlyGlyTyrGly                             355360365                                                                     GlyAlaGlyGlyGlyGluGlyValLeuMetGluGlyGlyValLysVal                              370375 380                                                                    ArgGluAlaValGluGluLysAsnLeuGlyGluAlaGlyArgGlyLeu                              385390395400                                                                  HisAlaCysAsnProAlaPheGlyArgProA rgGlnAlaAspTyrLeu                             405410415                                                                     ArgSerGlyValGlnAspGlnLeuGlyGlnGlnArgGluThrSerSer                              420 425430                                                                    LeuLeuLysIleGlnThrLeuAlaGlyHisGlyGlyArgArgLeu                                 435440445                                                                     (2) INFORMATION FOR SEQ ID NO:2:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 899 amino acids                                                    (B) TYPE: amino acid                                                         (D) TOPOLOGY: unknown                                                         (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:                                       MetGluSerCysTyrAsnProGlyLeuAspGlyIleIleGluTyrAsp                              1510 15                                                                       AspPheLysLeuAsnSerSerIleValGluProLysGluProAlaPro                              202530                                                                        GluThrAlaAspGlyProTyrLeuValIleValGluGln ProLysGln                             354045                                                                        ArgGlyPheArgPheArgTyrGlyCysGluGlyProSerHisGlyGly                              505560                                                                         LeuProGlyAlaSerSerGluLysGlyArgLysThrTyrProThrVal                             65707580                                                                      LysIleCysAsnTyrGluGlyProAlaLysIleGluValAs pLeuVal                             859095                                                                        ThrHisSerAspProProArgAlaHisAlaHisSerLeuValGlyLys                              100105 110                                                                    GlnCysSerGluLeuGlyIleCysAlaValSerValGlyProLysAsp                              115120125                                                                     MetThrAlaGlnPheAsnAsnLeuGlyValLeuHisVa lThrLysLys                             130135140                                                                     AsnMetMetGlyThrMetIleGlnLysLeuGlnArgGlnArgLeuArg                              145150155 160                                                                 SerArgProGlnGlyLeuThrGluAlaGluGlnArgGluLeuGluGln                              165170175                                                                     GluAlaLysGluLeuLysLysValMetAspL euSerIleValArgLeu                             180185190                                                                     ArgPheSerAlaPheLeuArgAlaSerAspGlySerPheSerLeuPro                              195200 205                                                                    LeuLysProValThrSerGlnProIleHisAspSerLysSerProGly                              210215220                                                                     AlaSerAsnLeuLysIleSerArgMetAspLysThr AlaGlySerVal                             225230235240                                                                  ArgGlyGlyAspGluValTyrLeuLeuCysAspLysValGlnLysAsp                              245 250255                                                                    AspIleGluValArgPheTyrGluAspAspGluAsnGlyTrpGlnAla                              260265270                                                                     PheGlyAspPheSerProThrAsp ValHisLysGlnTyrAlaIleVal                             275280285                                                                     PheArgThrProProTyrHisLysMetLysIleGluArgProValThr                              290295 300                                                                    ValPheLeuGlnLeuLysArgLysArgGlyGlyAspValSerAspSer                              305310315320                                                                  LysGlnPheThrTyrTyrProLe uValGluAspLysGluGluValGln                             325330335                                                                     ArgLysArgArgLysAlaLeuProThrPheSerGlnProPheGlyGly                              340 345350                                                                    GlySerHisMetGlyGlyGlySerGlyGlyAlaAlaGlyGlyTyrGly                              355360365                                                                     GlyAlaGlyGlyGlyGlyS erLeuGlyPhePheProSerSerLeuAla                             370375380                                                                     TyrSerProTyrGlnSerGlyAlaGlyProMetGlyCysTyrProGly                              385390 395400                                                                 GlyGlyGlyGlyAlaGlnMetAlaAlaThrValProSerArgAspSer                              405410415                                                                     GlyGluGluAla AlaGluProSerAlaProSerArgThrProGlnCys                             420425430                                                                     GluProGlnAlaProGluMetLeuGlnArgAlaArgGluTyrAsnAla                              43 5440445                                                                    ArgLeuPheGlyLeuAlaGlnArgSerAlaArgAlaLeuLeuAspTyr                              450455460                                                                     GlyValThrAlaAspAla ArgAlaLeuLeuAlaGlyGlnArgHisLeu                             465470475480                                                                  LeuThrAlaGlnAspGluAsnGlyAspThrProLeuHisLeuAlaIle                               485490495                                                                    IleHisGlyGlnThrSerValIleGluGlnIleValTyrValIleHis                              500505510                                                                     HisAl aGlnAspLeuGlyValValAsnLeuThrAsnHisLeuHisGln                             515520525                                                                     ThrProLeuHisLeuAlaValIleThrGlyGlnThrSerValValSer                               530535540                                                                    PheLeuLeuArgValGlyAlaAspProAlaLeuLeuAspArgHisGly                              545550555560                                                                  AspS erAlaMetHisLeuAlaLeuArgAlaGlyAlaGlyAlaProGlu                             565570575                                                                     LeuLeuArgAlaLeuLeuGlnSerGlyAlaProAlaValProGlnLeu                               580585590                                                                    LeuHisMetProAspPheGluGlyLeuTyrProValHisLeuAlaVal                              595600605                                                                      ArgAlaArgSerProGluCysLeuAspLeuLeuValAspSerGlyAla                             610615620                                                                     GluValGluAlaThrGluArgGlnGlyGlyArgThrAlaLeuHisLeu                              62 5630635640                                                                 AlaThrGluMetGluGluLeuGlyLeuValThrHisLeuValThrLys                              645650655                                                                     LeuArgAlaAsnValAsnAlaArgThrPheAlaGlyAsnThrProLeu                              660665670                                                                     HisLeuAlaAlaGlyLeuGlyTyrProThrLeuThrArgLeu LeuLeu                             675680685                                                                     LysAlaGlyAlaAspIleHisAlaGluAsnGluGluProLeuCysPro                              690695700                                                                      LeuProSerProProThrSerAspSerAspSerAspSerGluGlyPro                             705710715720                                                                  GluLysAspThrArgSerSerPheArgGlyHisThrProLeu AspLeu                             725730735                                                                     ThrCysSerThrLysValLysThrLeuLeuLeuAsnAlaAlaGlnAsn                              740745 750                                                                    ThrMetGluProProLeuThrProProSerProAlaGlyProGlyLeu                              755760765                                                                     SerLeuGlyAspThrAlaLeuGlnAsnLeuGluGlnLe uLeuAspGly                             770775780                                                                     ProGluAlaGlnGlySerTrpAlaGluLeuAlaGluArgLeuGlyLeu                              785790795 800                                                                 ArgSerLeuValAspThrTyrArgGlnThrThrSerProSerGlySer                              805810815                                                                     LeuLeuArgSerTyrGluLeuAlaGlyGlyA spLeuAlaGlyLeuLeu                             820825830                                                                     GluAlaLeuSerAspMetGlyLeuGluGluGlyValArgLeuLeuArg                              835840 845                                                                    GlyProGluThrArgAspLysLeuProSerThrGluValLysGluAsp                              850855860                                                                     SerAlaTyrGlySerGlnSerValGluGlnGluAla GluLysLeuGly                             865870875880                                                                  ProProProGluProProGlyGlyLeuCysHisGlyHisProGlnPro                              885 890895                                                                    GlnValHis                                                                     (2) INFORMATION FOR SEQ ID NO:3:                                              (i) SEQUENCE CHARACTERISTICS:                                                 (A) LENGTH: 969 amino acids                                                   (B) TYPE: amino acid                                                          (D) TOPOLOGY: unknown                                                         (ii) MOLECULE TYPE: protein                                                   (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3:                                       MetAlaGluAspAspProTyrLe uGlyArgProGluGlnMetPheHis                             151015                                                                        LeuAspProSerLeuThrHisThrIlePheAsnProGluValPheGln                              20 2530                                                                       ProGlnMetAlaLeuProThrAlaAspGlyProTyrLeuGlnIleLeu                              354045                                                                        GluGlnProLysGlnArgGly PheArgPheArgTyrValCysGluGly                             505560                                                                        ProSerHisGlyGlyLeuProGlyAlaSerSerGluLysAsnLysLys                              6570 7580                                                                     SerTyrProGlnValLysIleCysAsnTyrValGlyProAlaLysVal                              859095                                                                        IleValGlnLeuValTh rAsnGlyLysAsnIleHisLeuHisAlaHis                             100105110                                                                     SerLeuValGlyLysHisCysGluAspGlyIleCysThrValThrAla                              115 120125                                                                    GlyProLysAspMetValValGlyPheAlaAsnLeuGlyIleLeuHis                              130135140                                                                     ValThrLysLysLysValPheG luThrLeuGluAlaArgMetThrGlu                             145150155160                                                                  AlaCysIleArgGlyTyrAsnProGlyLeuLeuValHisProAspLeu                               165170175                                                                    AlaTyrLeuGlnAlaGluGlyGlyGlyAspArgGlnLeuGlyAspArg                              180185190                                                                     GluLysGlu LeuIleArgGlnAlaAlaLeuGlnGlnThrLysGluMet                             195200205                                                                     AspLeuSerValValArgLeuMetPheThrAlaPheLeuProAspSer                              210 215220                                                                    ThrGlySerPheThrArgArgLeuGluProValValSerAspAlaIle                              225230235240                                                                  TyrAspSer LysAlaProAsnAlaSerAsnLeuLysIleValArgMet                             245250255                                                                     AspArgThrAlaGlyCysValThrGlyGlyGluGluIleTyrLeuLeu                               260265270                                                                    CysAspLysValGlnLysAspAspIleGlnIleArgPheTyrGluGlu                              275280285                                                                     GluGl uAsnGlyGlyValTrpGluGlyPheGlyAspPheSerProThr                             290295300                                                                     AspValHisArgGlnPheAlaIleValPheLysThrProLysTyrLys                              305 310315320                                                                 AspIleAsnIleThrLysProAlaSerValPheValGlnLeuArgArg                              325330335                                                                      LysSerAspLeuGluThrSerGluProLysProPheLeuTyrTyrPro                             340345350                                                                     GluIleLysAspLysGluGluValGlnArgLysArgGlnLysLeuMe t                             355360365                                                                     ProAsnPheSerAspSerPheGlyGlyGlySerGlyAlaGlyAlaGly                              370375380                                                                     Gly GlyGlyMetPheGlySerGlyGlyGlyGlyGlyGlyThrGlySer                             385390395400                                                                  ThrGlyProGlyTyrSerPheProHisTyrGlyPheProThrTyrG ly                             405410415                                                                     GlyIleThrPheHisProGlyThrThrLysSerAsnAlaGlyMetLys                              420425 430                                                                    HisGlyThrMetAspThrGluSerLysLysAspProGluGlyCysAsp                              435440445                                                                     LysSerAspAspLysAsnThrValAsnLeuPheGlyLysVal IleGlu                             450455460                                                                     ThrThrGluGlnAspGlnGluProSerGluAlaThrValGlyAsnGly                              465470475 480                                                                 GluValThrLeuThrTyrAlaThrGlyThrLysGluGluSerAlaGly                              485490495                                                                     ValGlnAspAsnLeuPheLeuGluLysAlaMetGln LeuAlaLysArg                             500505510                                                                     HisAlaAsnAlaLeuPheAspTyrAlaValThrGlyAspValLysMet                              515520 525                                                                    LeuLeuAlaValGlnArgHisLeuThrAlaValGlnAspGluAsnGly                              530535540                                                                     AspSerValLeuHisLeuAlaIleIleHisLeuHisSerGl nLeuVal                             545550555560                                                                  ArgAspLeuLeuGluValThrSerGlyLeuIleSerAspAspIleIle                              565570 575                                                                    AsnMetArgAsnAspLeuTyrGlnThrProLeuHisLeuAlaValIle                              580585590                                                                     ThrLysGlnGluAspValValGluAspL euLeuArgAlaGlyAlaAsp                             595600605                                                                     LeuSerLeuLeuAspArgLeuGlyAsnSerValLeuHisLeuAlaAla                              610615 620                                                                    LysGluGlyHisAspLysValLeuSerIleLeuLeuLysHisLysLys                              625630635640                                                                  AlaAlaLeuLeuLeuAspHisProAsn GlyAspGlyLeuAsnAlaIle                             645650655                                                                     HisLeuAlaMetMetSerAsnSerLeuProCysLeuLeuLeuLeuVal                              660 665670                                                                    AlaAlaGlyAlaAspValAsnAlaGlnGluGlnLysSerGlyArgThr                              675680685                                                                     AlaLeuHisLeuAlaValGluHis AspAsnIleSerLeuAlaGlyCys                             690695700                                                                     LeuLeuLeuGluGlyAspAlaHisValAspSerThrThrTyrAspGly                              705710 715720                                                                 ThrThrProLeuHisIleAlaAlaGlyArgGlySerThrArgLeuAla                              725730735                                                                     AlaLeuLeuLysAlaAl aGlyAlaAspProLeuValGluAsnPheGlu                             740745750                                                                     ProLeuTyrAspLeuAspAspSerTrpGluAsnAlaGlyGluAspGlu                              755 760765                                                                    GlyValValProGlyThrThrProLeuAspMetAlaThrSerTrpGln                              770775780                                                                     ValPheAspIleLeuAsnGlyL ysProTyrGluProGluPheThrSer                             785790795800                                                                  AspAspLeuLeuAlaGlnGlyAspMetLysGlnLeuAlaGluAspVal                               805810815                                                                    LysLeuGlnLeuTyrLysLeuLeuGluIleProAspProAspLysAsn                              820825830                                                                     TrpAlaThr LeuAlaGlnLysLeuGlyLeuGlyIleLeuAsnAsnAla                             835840845                                                                     PheArgLeuSerProAlaProSerLysThrLeuMetAspAsnTyrGlu                              850 855860                                                                    ValSerGlyGlyThrValArgGluLeuValGluAlaLeuArgGlnMet                              865870875880                                                                  GlyTyrThr GluAlaIleGluValIleGlnAlaAlaSerSerProVal                             885890895                                                                     LysThrThrSerGlnAlaHisSerLeuProLeuSerProAlaSerThr                               900905910                                                                    ArgGlnGlnIleAspGluLeuArgAspSerAspSerValCysAspThr                              915920925                                                                     GlyVa lGluThrSerPheArgLysLeuSerPheThrGluSerLeuThr                             930935940                                                                     SerGlyAlaSerLeuLeuThrLeuAsnLysMetProHisAspTyrGly                              945 950955960                                                                 GlnGluGlyProLeuGluGlyLysIle                                                   965                                                                       

What is claimed as new and desired to be secured by Letters Patent ofthe United States is:
 1. An isolated DNA sequence which encodes theprotein of the sequence shown in SEQ ID NO.:1.
 2. An isolated RNAsequence which encodes the protein of the sequence shown in SEQ IDNO.:1.
 3. A cell transformed with the isolated DNA sequence of claim 1.4. An isolated DNA sequence which encodes the protein of the sequenceshown in SEQ ID NO.:2.
 5. An isolated RNA sequence which encodes theprotein of the sequence shown in SEQ ID NO.:2.
 6. A cell transformedwith the isolated DNA sequence of claim
 4. 7. An isolated antisense RNAsequence which is fully complementary to the RNA sequence of claim 2 or5.
 8. The DNA sequence of claim 1 further comprising a κB-containinggene.
 9. A cell comprising a κB-containing gene and the DNA sequence ofclaim 1, wherein said DNA sequence is capable of stimulating theexpression of said κB-containing gene.
 10. The cell of claim 9, whereinsaid cell is a prokaryotic cell.
 11. The cell of claim 9, wherein saidcell is a eukaryotic cell.
 12. The isolated DNA sequence of claim 4further comprising a κB-containing gene.
 13. A cell comprising aκB-containing gene and the isolated DNA sequence of claim 4, whereinsaid DNA sequence is capable of stimulating the expression of saidκB-containing gene.